Tip-enhanced single molecule fluorescence near-field microscopy in aqueous environment

For nanobiophysical applications, scanning near-field optical microscopy must combine high optical resolution and single fluorescent molecule sensitivity with the ability to operate in aqueous solution. These requirements can be achieved using the electric field enhancement at the tip of illuminated silicon probes for atomic force miscroscopy (AFM), whereby single ATTO-740 dye molecules could be imaged at an optical resolution down to 20 nm under ambient conditions as well as in aqueous solution. Two illumination modes have been tested: (a) bottom illumination in a total internal reflection microscopy setup and (b) direct top illumination, both with dedicated phase-sensitive single photon counting technology in dynamic AFM mode of operation.